The ability to grow by using a multitude of potential nutritional sources is a hallmark of the bacterial world. As a result, a goal of microbial physiology is obtaining an understanding of how microorganisms sense alterations in their environment, and how they respond rapidly and efficiently. Bacteria adapt readily and efficiently and have evolved very sensitive means of monitoring changes in their intracellular pools or in their immediate environments and highly sophisticated mechanisms for translating those signals into changes in gene expression.
Sporulation is a special case of adaptation to nutrient limitation. It is general in the sense that limitation of any of sources of carbon, nitrogen or phosphorus can induce sporulation, but the decision to sporulate in no way ameliorates the stress-inducing condition (Sonenshein, A. L., 2000. In: Bacterial stress responses, G. Storz et al. (eds.), ASM Press, Washington, D.C., pp. 199-215). Instead the sporulating cell gives up on growth in order to create within itself a protected environment for its genome. While the spore is highly resistant to many forms of environmental stress (e.g., heat, desiccation, large variations in pH, organic solvents, osmotic imbalance, antibiotics), spore formation is not triggered by any of the stresses to which resistance is gained. The specific metabolic signal to which cells respond when making the decision to sporulate and the mechanism by which the signal is recognized remain incompletely characterized (Sonenshein, A. L., 1989. In: Regulation of procaryotic development, I. Smith et al. (eds.), ASM Press, Washington, D.C., pp. 109-130). Freese and colleagues showed that a drop in the intracellular pools of GDP and GTP correlates with the onset of sporulation (Freese, E. et al., 1979. J. Gen. Microbiol. 115:193-205; Lopez, J. et al., 1981. J. Bacteriol. 146:605-61; Lopez, J. et al., 1979. Biochim Biophys. Acta. 587:238-252; Mitani, T. et al., 1977. Biochim. Biophys. Acta. 77:1118-1125; Ochi, K. et al., 1981. J. Biol. Chem. 256:6866-6875). Moreover, limitation of guanine nucleotide synthesis, by treatment with an inhibitor or by limitation of a purine auxotroph, induces sporulation in cultures growing in excess nutrients (Freese, E. et al., 1979. J. Gen. Microbiol. 115:193-205; Freese, E. et al., 1979. Mol. Gen. Genet. 170:67-74; Mitani, T. et al., 1977. Biochim. Biophys. Acta. 77:1118-1125), as if the nutritional signal that cells monitor is the concentration of GDP and/or GTP. Nucleotides are exemplary as signal compounds because their synthesis depends on adequate supplies of carbon, nitrogen and phosphorus.
For many years researchers have sought the B. subtilis proteins that sense nutrient limitation and, more specifically, GTP availability. Recent evidence indicates that CodY, a GTP binding protein, is a factor in this regulation (Ratnayake-Lecamwasam, M. et al., 2001. Genes Dev. 15:1093-1103). In fact, GTP activates CodY as a repressor (Ratnayake-Lecamwasam, M. et al., 2001). CodY was first identified as a negative regulator of the dipeptide permease (dpp) operon (Slack, F. et al., 1993. J. Bacteriol. 175:4605-4614; Slack, F. et al., 1995. Mol. Microbiol. 15:689-702), and was subsequently found to repress during rapid exponential growth phase a group of genes whose products are generally involved in adaptation to poor growth conditions. The targets of CodY include genes that encode extracellular degradative enzymes, transport systems, catabolic pathways, genetic competence, antibiotic synthesis, flagellin, and sporulation functions (Burkholder, W. et al., 2000. In: Prokaryotic development, Y. Brun et al. (eds.), ASM Press, Washington D.C., pp. 151-166; Debarbouille, M. et al., 1999. J. Bacteriol. 181:2059-2066; Ferson, A. et al., 1996. Mol. Microbiol. 22:693-701; Fisher, S. et al., 1996. J. Bacteriol. 178:3779-3784; Mirel, D. et al., 2000. J. Bacteriol. 182:3055-3062; Serror, P. et al., 1996. J. Bacteriol. 178:5910-5915; Slack, F. et al., 1993. J. Bacteriol. 175:4605-4614; Slack, F. et al., 1995. Mol. Microbiol. 15:689-702; Wray, L. et al., 1997. J. Bacteriol. 179:5494-5501; Molle, V. et al., 2003. J. Bacteriol. 185:1911-1922; Kim, H. et al., 2003. J. Bacteriol. 185:1672-1680).
Although unrelated to other families of regulatory proteins, homologs of CodY are generally found in the low G+C family of Gram positive bacteria, including Bacillus anthracis, B. halodurans, B. stearothermophilus, Clostridium perfringens, C. difficile, C. acetobutylicum, C. botulinum, Staphylococcus aureus, S. epidermidis, Streptococcus pneumoniae, S. agalactiae, S. pyogenes, S. equi, S. mutans, Listeria monocytogenes, L. innocua, Desulfitobacterium hafniense, Carboxydothermus hydrogenoformans, Lactococcus lactis, and Enterococcus faecalis. This group includes major human and animal pathogens and important industrial bacteria.
With recent increases in antibiotic resistance in bacteria, there is a need for compositions and methods for regulating production of toxins and other materials involved in disease by Gram positive pathogens.